Cytochromes P-450, a class of cellular hemoprotein enzymes, act as monooxygenases in catalyzing metabolic functions. It fluctuates between ferrous and ferric oxidation states in acting as an electron transporter. It will catalyze the insertion of an oxygen atom into C--H bonds of organic compounds using dissolved molecular oxygen. The oxygen is cleaved, one atom inserting itself into the C--H bond and the other forming a molecule of water with hydrogen from NADPH and a reductase. The presence of iron in the cytochrome 450 renders the oxygen highly reactive.
The process is, however, limited by the availability of the protein cytochrome P-450, by the need for the presence of a reductase, and by the stoichiometric involvement of NADPH (nicotinamide-adenine dinucleotide phosphate reduced) as the reducing agent. The protein availability problem can be overcome, at least in theory, by the substitution for cytochrome P-450 of another, more readily available protein capable of exhibiting similar monooxygenase activity.
It has previously been reported that methemoglobin, a very abundant protein formed by the oxidation of hemoglobin to its ferric state, can substitute, in such C--H--C--OH bond conversions, for cytochrome P-450 in a system which utilizes NADPH as an electron source and cytochrome P-450 reductase to mediate electron transfer (Myeal et al, J. Biol. Chem. 1976, 251, p. 3436).
It has further been reported that a flavin-hemoglobin coupled product will function similarly and efficiently, without a reductase (Kokubo, et al, J. Am. Chem. Soc., 1987, 109, p. 606)). However, the utility of both of these systems is limited by the need for the presence of NADPH, and further, in the latter case, by the complexity of the preparation of the flavin-hemoglobin coupled product.
It is an object of the present invention to provide a novel, heme protein-catalyzed process for conducting C--H--C--OH bond conversions in organic compounds having affinity for globin proteins.
It is a further object of the invention to provide such a process which does not depend on the presence in the system of the reductase and NADPH.